It has been generally known that allergic reactions such as asthma, rhinitis, urticaria, anaphylaxis, allergic bronchiectasis, allergy caused by food•drug•pollen•bug, hay fever, cold urticaria, and atopic dermatitis, are attributed to unbalance between cytokine involved in the regulation of IgE (Immunoglobulin E) secretion and IgE (Immunoglobulin E) over-expressed by hypersensitive reaction against allergen (Bachert et al., Clinical and Experimental Allergy 28,15-19, 1998; MacDonald and Lichtenstein, Springer Semin Immunopathol., 12, 415-428, 1990).
Once exposed on allergen, immediate reaction occurs, and then cells involved in inflammation are gathered around allergen exposed area by the action of cytokine secreted in mast cells. Few hours later, late-phase reaction (referred as “LPR” hereinafter) occurs owing to various cytokines and histamine secreted from basophils, eosinophils and lymphocytes, and the LPR is progressed in half of allergy patients. Under LPR, histamine is secreted from basophils. And in that case, there is No: allergen causing immediate reaction, so searching for a cause of histamine secretion and progress to LPR has been a major concern. It has been known that histamine is secreted by cytokines such as MCP-3 (Monocyte Chemotatic Protein-3), MCP-1 (Monocyte Chemotatic Protein-1) or RANTES (Regulated upon Activation Normal T-cell Expressed and Secreted), but in fact, only the protein named “HRF” has been proved to secrete histamine in basophils under IgE-dependent LPR (MacDonald et al., Science, 269, 688-690,1995). Nevertheless, the mechanism of HRF inducing secretion of histamine in basophils has not been disclosed yet.
HRF is a well-known protein composed of 172 amino acids, and is found in every cytoplasm (Bohm et al., Biochem. Int., 19, 277-286,1989). Among these HRF forming amino acids, 79-123 amino acids form a basic domain, which shows 46% homology with MAP-1B (microtubule-associated protein-1B). Thus, it is presumed that the protein is capable of binding to microtubule. Gachet et al observed under confocal microscope that HRF distribution is consistent with cytoskeleton network distribution, suggesting that HRF is bound to cytoskeleton (Gachet et al., J. Cell Sci., 112,1257-1271, 1999). In the meantime, Sanchez et al reported that HRF is bound to Ca2+ although HRF does not belong to calcium binding protein family. It was additionally reported that yeast could survive even when HRF gene was defected in Saccaromyces cerevisiae (Sanchez et al., Electrophoresis, 18, 150-155, 1997). The reports indicate that HRF belongs to a gene family which has redundant pathway.
HRF is a hydrophilic protein residing in cytoplasm, but has been found in outside of cytoplasm by MacDonald et al. Besides, high level of HRF is detected in serum of a LPR patient, indicating that HRF is extracellular secreted by apoptosis or any other mechanism so as to release histamine through HRF receptors residing on cell membrane (MacDonald et al., Science, 269, 688-690, 1995). HRF has known to stimulate IgE-sensitized basophils to secrete histamine, yet concrete IgE mechanism involved in the secretion has not been explained. According to Bheekha-Escura et al, HRF might cause inflammatory reaction in IgE receptor defected cells, suggesting that HRF is not bound directly to IgE but bound to a specific cell membrane receptor (Bheekha-Escura et al., Blood, 96, 2191-2198, 2000).
The present inventors have previously reported that i) HRF can pass through cell membrane even though it is a hydrophilic protein, and ii) HRF receptor is confirmed by yeast two-hybrid assay to be the third cytoplasmic domain (CD3) of (Na,K)ATPase (Korean Registered Patent Nos. 457350B1 and 457351B1). The inventors also explained the histamine releasing mechanism of HRF in basophils.
The present inventors found peptide which i) intercepts HRF's passing through the cell and/or ii) intercepts HRF's binding to (Na, K)ATPase to inhibit histamine secretion, and confirmed that such peptide can be useful for the prevention or the treatment of allergic diseases (Korean Registered Patent No. 457350B1).
According to recent report by Budde et al, histamine releasing activity of HRF (HRFmn) isolated from activated monocyte culture supernatant is different from that of recombinant HRF (rHRF), which suggests that HRFmn is not identical factor to rHRF (Budde et al., Ann. Allergy Asthma Immunol., 89, 606-612, 2002). In addition, the report also mentioned that HRF was not detected in HRFmn from human HRF specific ELISA (enzyme-linked immunoabsorbant assay), which suggests that HRFmn has a different structure from rHRF.
Under the judgment that the difference in activity between HRFmn and rHRF is attributed to protease which is massively distributed in allergy patient's blood, the present inventors tried to separate HRF having histamine releasing activity, and as a result, the present inventors found out deletion forms of HRF having better activity than wild type HRF. These deletion forms of HRF, unlike wild type HRF, have intermolecular disulfide bond, by which dimerization is induced and HRF is activated accordingly. The present inventors also prepared novel HRF-binding peptides, and confirmed that the bond of novel HRF-binding peptides to the deletion forms of HRF results in the inhibition of histamine and IL-8 secretion. The present inventors also confirmed that modified HRF binding peptides can be bound to deletion forms of HRF to inhibit histamine and IL-8 releasing activity of HRF, and thus completed this invention by further confirming that deletion forms of HRF and HRF binding peptides can be effectively used for the development of an anti-allergy drug.